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Method for screening new internal reference molecules suitable for cervical tissue micro RNA real-time fluorescence quantitative PCR research

A technology for real-time fluorescence quantification and screening methods, applied in fluorescence/phosphorescence, biochemical equipment and methods, material excitation analysis, etc., it can solve the problem of unexplored constancy of expression and the applicability of internal reference, and improve accuracy and reliability. The effect of stability, good calibration and normalization

Inactive Publication Date: 2010-09-01
ATTACHED OBSTETRICS & GYNECOLOGY OSPITAL MEDICALCOLLEGE ZHEJIANG UNIV
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Problems solved by technology

[0006] So far, the quantitative research on specific micro RNA related to cervical lesions has mostly blindly referred to internal reference molecules (RNU48, 5S, let-7a) used in other tumors, but the constant expression in cervical tissue Neither the sex nor its applicability as an internal control were explored

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  • Method for screening new internal reference molecules suitable for cervical tissue micro RNA real-time fluorescence quantitative PCR research
  • Method for screening new internal reference molecules suitable for cervical tissue micro RNA real-time fluorescence quantitative PCR research
  • Method for screening new internal reference molecules suitable for cervical tissue micro RNA real-time fluorescence quantitative PCR research

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Embodiment Construction

[0027] Below in conjunction with accompanying drawing, the present invention is further explained and illustrated:

[0028] Such as Figure 1-8 Shown:

[0029] The present invention selects 30 cases of normal cervical epithelial tissues and 30 cervical cancer tissues. The source of normal cervical epithelium is collected from volunteers who come to the Obstetrics and Gynecology Hospital Affiliated to Zhejiang University School of Medicine for cervical disease screening. Cervical cancer patients treated (23 cases of squamous cell carcinoma, 4 cases of adenosquamous cell carcinoma, 3 cases of adenocarcinoma). All specimens were biopsied tissues under colposcopy, and each specimen was divided into two parts, one was quick-frozen in liquid nitrogen and then stored in liquid nitrogen, and the other was fixed in 4% formaldehyde and embedded in stone vinegar. Liquid nitrogen cryopreserved tissue was used for Micro RNA chip inspection to screen internal reference molecules and to ve...

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Abstract

The invention discloses a method for screening new internal reference molecules suitable for cervical tissue micro RNA real-time fluorescence quantitative PCR research. The method comprises the following technical steps: screening human Micro RNA probes by using a Micro RNA chip, and selecting Micro RNAs with constant normal tissue expression and high expression abundance of cervical cancer and cervix as candidate internal references; selecting a normal cervical epithelial tissue and a cancerous cervical tissue as clinical cervical tissue specimens respectively, and verifying the specimens by using fluorescence quantitative PCR; and analyzing data by using two special internal reference analysis software comprising geNorm software and Norm Finder so as to discover the internal reference molecules suitable for cervical tissue Micro RNA research. The internal reference molecules screened and verified by using the method of the invention can avoid possible differences of different cervical tissue specimens on the yield, quality and reverse transcription efficiency of the RNA, well correct and standardize the data for real-time fluorescence quantitative PCR detection, and improve the accuracy and the reliability of the search.

Description

technical field [0001] The invention relates to a method for screening new internal reference molecules suitable for real-time fluorescent quantitative PCR research on micro RNA in cervical tissue. Background technique [0002] Cervical cancer is the second most common malignant tumor in women after breast cancer. A large number of epidemiological data show that high-risk human papillomavirus (HPV) infection is a necessary condition for the onset of cervical cancer and its precancerous lesions. But a single HPV infection is not enough to cause malignant transformation of cervical epithelium. Micro RNA (miRNA, referred to as miRNA) is a small non-coding RNA with a length of about 20-23 nucleotides in an organism, which negatively regulates gene expression at the post-transcriptional level through complementary pairing with target mRNA. Causes mRNA degradation or translational repression. So far, thousands of miRNAs have been reported to exist in multicellular eukaryotes suc...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N21/64G06F19/00
Inventor 谢幸沈源明吕卫国叶枫李阳王芬芬万小云
Owner ATTACHED OBSTETRICS & GYNECOLOGY OSPITAL MEDICALCOLLEGE ZHEJIANG UNIV
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